The receptor tyrosine kinases (RTKs) of the TAM family - Tyro3, Axl, and Mer - are exploited by flaviviruses, such as West Nile Virus and Dengue Virus, to infect their target cells. The experiments of this proposal are designed to elucidate how TAM potentiation of infection is achieved. TAM RTKs are expressed on the surface of many cells, including dendritic cells and macrophages of the immune system, and neurons of the central nervous system. The ligands that bind to and activate the TAMs - Gas6 and Protein S (ProS) - have the ability to attach both to the phospholipid phosphatidylserine, expressed on the surface of flavivirus particles, and also to their cognate TAM receptor. In this way, the TAM ligands serve as a 'bridge' that links flaviviruses to the surface of cells that they will infect. t the same time, TAM receptor activation by Gas6 or ProS results in the inhibition of type I interferon (IFN) signaling, to which flaviviruses and many other viruses are sensitive. In Aim 1, genetic, biochemical, and cell biological methods will be used to dissect the mechanisms through which TAM receptors facilitate flavivirus entry into human cells in vitro. In Aim 2, these same methods will be used in experiments in dendritic cells and macrophages prepared from mice that carry targeted mutations in the Axl and Mer genes, and in CNS neurons prepared Tyro3 mutant mice, to define the TAM receptors and ligands that promote infection in different cellular settings, and to determine the extent to which TAM facilitation of flavivirus infection is dependent on the inhibition of type I IFN signaling. In Aim 3, mice that carry mutations in the Tyro3, Axl, Mer, Gas6, and ProS genes will be used to assess the cellular and immunological consequences of specific deficits in TAM signaling for infection by West Nile Virus in vivo. Together, these experiments will delineate the molecular, cellular, and physiological features of a heretofore unknown pathway of flavivirus infection. PUBLIC HEALTH RELEVANCE: Each year, ~100 million people are infected by West Nile Virus, Dengue Virus, and related flaviviruses, which are the most important arthropod-borne viruses causing disease in humans. As there is no specific therapy available for the majority of these infections, it is essential to public health to understand fully the genetic, cell biological and biochemical mechanisms that underlie the potentiation of flavivirus infection by receptor tyrosine kinases of the TAM family.